|
| Status |
Public on Jun 04, 2014 |
| Title |
mEpiSC_H3K27me3 |
| Sample type |
SRA |
| |
|
| Source name |
Epiblast stem cells
|
| Organism |
Mus musculus |
| Characteristics |
cell line: EpiSC9
chip antibody: H3K27me3 (Abcam #6002)
|
| Growth protocol |
mEpiSCs were cultured in Knockout DMEM supplemented with 20% KSR, 10ng/ml FGF2 (R&D Systems), 2mM Glutamax, 1x non-essential amino acids, and 0.1mM 2-mercaptoethanol and passaged every third day using 1.5 mg/ml collagenase type IV (Invitrogen) and trituration into small clumps of ~10-100 cells. Irradiated mouse embryonic fibroblasts (MEFs) served as a feeder layer for all pluripotent cell types and were maintained with DMEM supplemented with 10% fetal bovine serum, 2 mM glutamax, 1x non-essential amino acids, and 0.1 mM 2- mercaptoethanol. All cells were grown on Nunclon Δ-treated dishes or multiwell plates (Fisher Scientific) coated for 2 hours at 37°C with 0.1% (w/v) porcine gelatin (Sigma).
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
ChIPs were performed from 5x10^6 crosslinked mEpiSCs as previously described (Schmidt et al., 2009). The following antibodies were used for ChIP: rabbit anti-H3K4me1 (Abcam #8895), rabbit anti- H3K4me3 (Active Motif #39159), rabbit anti-H3K27ac (Abcam #4729), mouse anti-H3K27me3 (Abcam #6002).
Sequencing libraries were prepared as previously described (Schmidt et al., 2009).
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina Genome Analyzer II |
| |
|
| Data processing |
The FASTX-Toolkit (http://hannonlab.cshl.edu/fastx_toolkit/) was used to remove adapter sequences and trim read ends using a quality score cutoff of 20.
ChIP-seq data were aligned to mm9 genome assemblies (retrieved from http://cufflinks.cbcb.umd.edu/igenomes.html), using Bowtie v0.12.9 (Langmead et al., 2009), allowing reads with ≤ 2 mismatches and discarding reads with > 1 reportable alignment ("-m 1" parameter).
Peaks were detected with MACS v1.4 (Zhang et al., 2008), using an aligned input DNA sample as control.
Genome_build: mm9
Supplementary_files_format_and_content: bigwig files - Wiggle tracks stepped at 25 bp were generated by MACS, normalized to the median signal by chromosome and converted to bigwig format using the UCSC Genome Browser’s wigToBigWig utility (retrieved from http://hgdownload.cse.ucsc.edu/admin/exe/). bed files - MACS called peaks.
|
| |
|
| Submission date |
May 07, 2014 |
| Last update date |
May 15, 2019 |
| Contact name |
Paul J. Tesar |
| E-mail(s) |
paul.tesar@case.edu
|
| Organization name |
Case Western Reserve University School of Medicine
|
| Department |
Genetics and Genome Sciences
|
| Street address |
10900 Euclid Ave
|
| City |
Clev |
| State/province |
OH |
| ZIP/Postal code |
44106 |
| Country |
USA |
| |
|
| Platform ID |
GPL9250 |
| Series (2) |
| GSE57407 |
Epigenomic comparison of distinct pluripotent stem cell states reveals a new class of enhancers with roles throughout mammalian development (ChIP-seq) |
| GSE57409 |
Epigenomic comparison of distinct pluripotent stem cell states reveals a new class of enhancers with roles throughout mammalian development |
|
| Relations |
| BioSample |
SAMN02767303 |
| SRA |
SRX535241 |
| Named Annotation |
GSM1382220_mEpiSC_H3K27me3.bigWig |
